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Biomedical subjects

H B Benestad

Publications and source records attributed to H B Benestad.

At least 19 recordsLinked to original sources
PubMed1992

Blood flow to bone marrow during development of anemia or polycythemia in the rat.

We applied the radioactive microsphere method to follow the magnitude and time course (0 to 96 hours) of blood flow changes during development and recovery from anemia in awake rats. Blood flow was also monitored during a 96-hour period after polycythemia was induced (erythropoietin administered subcutaneously [SC]). The possible influence of innervation was also examined. After a blood loss of approximately 50% (hypovolemia), blood flow to the femoral marrow tripled within 12 hours and remained elevated for the entire 96-hour period. The relative increase in blood flow to the femoral bone was even greater. Similar findings were obtained in rats with phenylhydrazine (PHZ) hemolytic anemia (normovolemia). Denervation had no detectable effect on the increased blood flow to either marrow or bone. The augmented blood flow during hemolytic anemia was accompanied by a doubling of the oxygen consumption rate by the marrow, while the glucose uptake was not detectably altered. Erythropoietin supplements (3 x 1,000 IU/kg, SC, 6-hour intervals) increased blood flow to the marrow by approximately 25% after 48 hours, and at 72 hours the blood flow had reached a value twice that obtained under control conditions. These results indicate that blood flow to bone marrow is highly variable and hormonally and/or locally regulated. This may have practical consequences for marrow transplantation technology and for administration of drug therapy to patients with insufficient bone marrow hematopoiesis.

Anemia↗
PubMed1992

In vivo measurement of the association constant of a radio-labelled monoclonal antibody in experimental immunotargeting.

Exploring the fundamental mechanisms behind the low tumour uptake of labelled monoclonal antibodies (MoAbs) during in vivo immunotargeting, experiments were performed to estimate the in vivo value of the association constant (Ka) in an experimental targeting reaction. An artificial tumour model was utilised, based on diffusion chambers (DC) filled with antigen-coated polymer particles, implanted i.p. in normal, immunocompetent mice (NMRI/BOM). The MoAb H7 with specificity for placental alkaline phosphatase (PLALP) was chosen for this experiment. Each mouse carried two DC, one target DC filled with PLALP-coated particles, and a second control DC with the same amount of uncoated particles. The DC contained escalating doses of particles, ranging from 0.1 mg to 16 mg per DC, with groups of 6-12 animals per dose level. The next day after the implantation, a constant dose of 125I-labelled Fab fragments of H7 was injected i.v. in each mouse. The association constant Ka as measured from the binding data obtained in vivo was not significantly different from the value measured in vitro when the same target DC were incubated with the 125I-Fab in test tubes. This indicates that in vivo impairment of the antibody avidity is not the reason why a relatively low tumour uptake is generally experienced in immunotargeting studies.

Alkaline Phosphatase↗
PubMed1992

Marked splenic hyperaemia during post-haemorrhagic hypotension in the rat, rabbit and cat.

1. The regulation of the splenic perfusion during normal and pathological conditions is incompletely understood. We studied the time course of splenic blood flow during the initial (0-24 h) development of haemorrhagic anaemia in awake and anaesthetized rats, as well as in awake rabbits and cats. Another group of rats had either normovolaemic anaemia or beginning polycythaemia. 2. The microsphere method was used to measure splenic blood flow. The awake rats were rendered anaemic either by a heavy (1.5% of body weight) or a moderate (0.7% of body weight) bleeding (hypovolaemia), by haemolysis (normovolaemia) or by bleeding (1.5% of body weight) followed by transfusion of autologous plasma (normovolaemia). Polycythaemia was induced with injections of erythropoietin. The anaesthetized rats as well as the awake rabbits and cats were also bled heavily (1.5% of body weight). 3. In awake rats, splenic blood flow increased to 215% of control within the first 5 min after bleeding. The perfusion declined nearly to baseline over the next 24 h. A similar, but less prominent splenic hyperaemia was detected in the awake rabbits and cats. However, this hyperaemic response was not detected in the normovolaemic, the polycythaemic or the anaesthetized and bled rats. 4. Administration of the adrenergic beta-blocking agent propranolol prior to bleeding significantly attenuated the splenic hyperaemia in the awake rats, while the alpha-blocking agent phentolamine or the cholinergic blocking agent atropine had no effect. 5. Concomitant with the initial increase in splenic perfusion, cardiac output increased to 123% of control in the awake, heavily bled rats. In the bled, anaesthetized rats cardiac output decreased to 91% of control 5 min after bleeding. A decrease in cardiac output to 64 and 70% of control was observed in the awake rabbits and cats, respectively. 6. Immediately following the bleeding, we noticed a substantial release of platelets from the rat spleen. 7. It appears that a heavy acute blood loss in awake rats, rabbits and cats elicits a marked reduction in splenic vascular tone, perhaps mediated by beta-adrenergic receptor activity. Anaesthesia abolished this response in rats. Possibly, the induced hypovolaemia triggered an accelerated release of platelets from the rat spleen, dependent on an augmented splenic blood flow.

Anemia↗
PubMed1991

Pulmonary, hepatic and splenic sequestration of technetium-99m labelled autologous rabbit granulocytes: scintigraphic cell distributions after intravenous and intraarterial injections, exsanguination and intraarterial injection of cells passed through an intermediary host.

In man, about half the intravascular granulocytes are not freely circulating, but temporarily sequestered ('marginated'), so that they cannot be retrieved by bleeding. Where and how the sequestration occurs is not settled and is the subject of the present report. Isolated autologous rabbit granulocytes, labelled with two different 99mTc methods, were reinjected and followed with external scintigraphy. Intraarterial as well as intravenous injection led to rapid accumulation of radioactivity over the lungs. This finding was corroborated and extended by similar experiments, where the labelled cells had firstly been passed through an intermediary rabbit host to remove altered cells, i.e. cells damaged, 'primed' (pre-activated), or activated. In the final autologous host about two thirds of the label rapidly localized to the lungs and liver, and a few per cent to the spleen (which is very small in the rabbit). Even though more than half of the intermediary rabbit's calculated blood volume was removed, the blood sample contained only a few per cent of the rabbit's radioactivity; consequently, many of the labelled leucocytes had marginated during the bleeding. The proportional distribution of radioactivity over lungs, spleen, kidneys, and the rest of the intermediary animal was not markedly changed by this exsanguination, but there was a 4-20% decrease over the liver. Taken together, our findings indicate that normal granulocytes marginate in lungs, liver, and spleen--apparently explicable by the effects of cell size, vessel diameter, cell stiffness (visco-elastic properties) and size of the arterio-venous hydrostatic pressure difference. The liver and spleen seemed to play additional roles, since radioactivity over these organs decreased much slower than expected from reported blood half-times of intact and slightly damaged rabbit granulocytes. This led to a suggestion that macrophages exposed to blood normally phagocytose apoptotically dying granulocytes.

Animals↗
PubMed1991

Spontaneous alloreactivity of natural killer (NK) and lymphokine-activated killer (LAK) cells from athymic rats against normal haemic cells. NK cells stimulate syngeneic but inhibit allogeneic haemopoiesis.

We wanted to re-examine the hypotheses that natural killer (NK) cells preferentially react with immature cells, and that they are not directed against major histocompatibility complex (MHC) gene products. Rat marrow cells could be separated according to maturity on a four-step discontinuous density gradient of Percoll. Almost all the immature bone marrow cells with progenitor activity, as measured in vivo in a diffusion chamber assay or in vitro in a granulocyte/macrophage colony-forming assay, resided within the lighter density cell fraction (density approximately 1.065). The higher density cells (density approximately 1.082) contained mainly the more mature, non-proliferative cells within the granulocyte series. NK and lymphokine-activated killer (LAK) cells from athymic rats, being devoid of T cells, efficiently killed low- as well as high-density bone marrow cells from a fully allogeneic and a MHC congenic rat strain, while little or no killing was observed against syngeneic bone marrow cell fractions. LAK cells also effectively inhibited granulocyte/macrophage colony formation from allogeneic bone marrow precursors in vitro, while stimulating colony formation from syngeneic bone marrow cells. The NK-mediated killing of allogeneic bone marrow cells was effectively inhibited by NK-sensitive tumour cells, while there was much less inhibition of the killing of tumour cells by allogeneic bone marrow cells. We conclude that NK cells recognize MHC incompatibilities on both immature and mature allogeneic bone marrow cells through recognition systems not related to T-cell receptors, and that allospecific killing can explain the contrasting effect of NK cells on allogeneic and syngeneic haematopoiesis.

Animals↗
PubMed1990

Migration of 99mTc-labelled syngeneic lymphocytes in the rat. Biological and theoretical models predict radiation damage and poor scintigraphic detectability.

The possibility of obtaining useful scintigrams of secondary lymphoid organs after infusion of syngeneic lymphocytes labelled with technetium-99m (99mTc) was explored in a rat model. Thoracic duct lymphocyte (TDL) accumulation in various organs was measured with both 99mTc and 51Cr labelled cells, the latter processed with a method that has been shown not to damage lymphocytes. 99mTc labelled TDL did not localize properly in the lymph nodes and spleen. We could not visualize lymph nodes in scintigrams, neither could we demonstrate any difference between normal and hyperplastic spleens. Our conclusion is that radiation from the 99mTc label readily influences lymphocyte migration so that useful scintigraphy in rats and other small experimental animals becomes impossible. This was supported by results from culture experiments with 99mTc labelled, radiosensitive mouse haemopoietic progenitor cells. Theoretical considerations, including the calculations of lymphocyte self-irradiation and signal/noise ratios during scintigraphy of rat tissues, supported our conclusion that scintigraphy in small animals, to disclose the physiological migration of lymphocytes, may be impossible with the present sensitivity of gamma cameras.

Animals↗
PubMed1990

Radioimmunotargeting of human tumour cells in immunocompetent animals.

A tumour model system is reported that for many purposes may be an alternative to xenografted nude mice. The model allows immunotargeting of human tumour cells in immunocompetent animals. The target cells are contained in i.p. diffusion chambers (DC) with micropore membrane walls that are permeable to molecules, including the cell specific monoclonal antibodies (MoAb), but impermeable to cells. Thus, the tumour cells are protected from the host immunocompetent cells. In the work here presented the model was tested in immunocompetent mice and pigs, with tumour cells and antibody preparations that had demonstrated specific targeting in the nude mouse xenograft model. Hence, the DC were filled with cells from the human cell lines Hep-2 (expressing placental alkaline phosphatase, PLALP), or OHS (a sarcoma cell line), and the MoAb preparations injected i.v. were a 125I-labelled Fab fragment of the PLALP specific antibody H7, or a 125I-labelled F(ab')2 fragment of the sarcoma specific antibody TP-1. Specific targeting of the human tumour cells was demonstrated in both mice and pigs. The target: blood ratios were comparable in the two species, reaching a maximum of about 15 after 24 h with the Fab preparation, and a ratio of 25 after 72 h with the F(ab')2. The target uptake relative to injected dose was lower in pigs than in mice, but the difference between the two species was smaller than expected, presumably due to a slower antibody clearance in the pigs than in the mice. An artificial cell targeting system like this has several advantages in the search for solutions to many of the fundamental problems experienced in immunotargeting. Firstly, parallel binding experiments can be carried out in vitro with the same target. Because in vitro results are only influenced by the diffusion into the DC and the immunological binding characteristics of the antibodies, targeting differences between antibody preparations due to these factors can then be distinguished from differences due to pharmacokinetical properties. Secondly, the animals can be implanted with any type and number of target cells, or with antigen negative control cells. Thirdly, and perhaps most important, the system opens a possibility for evaluation of the murine MoAb in xenogenic species, and this may predict the clinical targeting potential better than experiments on mice.

Animals↗
PubMed1990

Functional capacity of neutrophil granulocytes in deep-sea divers.

Neutrophil granulocytes (PMN) are main defenders against invading microbes. We evaluated the adaptive response of PMN from divers exposed for weeks to high total and oxygen pressures. Under these conditions PMN could be primed to give a heightened respiratory burst upon stimulation with the bacterial peptide analogue, formyl-methionyl-leucyl-phenylalanine (FMLP): blood PMN sampled both shortly after operational saturation dives offshore and during an onshore test-dive gave larger responses than control pre- or post-dive PMN from the same subjects and PMN from laboratory personnel. The assays used measured oxygen consumption, intracellular H2O2 availability, and chemiluminescence. The submaximal responses provoked by the non-metabolizable diacylglycerol analogue phorbol myristate acetate (PMA) were less and less often increased. Such enhanced PMN responsiveness may possibly decrease resistance to skin and other infections that are encountered in divers, if PMN thereby failed to localize correctly to inflamed tissues.

Adult↗
PubMed1989

[Increased importance of teaching qualifications? Reality or only empty words?].

Educational qualifications are important for all teachers of medicine. At University of Oslo, Norway, yearly personnel development courses are held for newly appointed teaching faculty members. To find out what priority the Medical School gives to this course, and how it is evaluated by the participants in the medical course, we conducted a questionnaire survey among the participants. The course was very well received by the participants, but the results indicate that personnel development is given low priority by the Medical School.

Education, Medical, Continuing↗
PubMed1989

Leucopoiesis versus concentration of cytokines in diffusion chamber cultures of mouse bone marrow cells: clues to the physiological roles of growth factors.

Physiological mechanisms that regulate formation of neutrophil granulocytes, macrophages and their precursor cells were studied with the diffusion chamber (DC) technique. DC inoculated with mouse bone marrow cells were implanted intraperitoneally into host mice. When these in vivo cultures had been established and their marrow populations were expanding (2-day cultures), the DC were transferred to different environments: new, normal mice, lethally irradiated mice, or incubation flasks with optimal concentrations of growth factors. Culture development during the following final culture period was correlated to the concentration of some select candidate growth regulators in DC. After 3 d the cellularity of DC in irradiated hosts had increased significantly more than in the normal hosts. A difference was detectable already after 1-2 d when preculturing was omitted. The increased growth appeared to take place at several stages of cell maturation and not only at the progenitor cell level. Colony stimulating factor(s) for granulocyte and macrophage progenitors, as well as cytokines stimulating the bone marrow-derived cell line, 14 M.1, were present in DC fluid (DCF) at higher concentrations in irradiated than in normal mice throughout the final culture period. On the other hand, DCF concentrations of tumour necrosis-like factor (that may either induce CSF secretion or directly inhibit myelopoiesis), were not significantly different in irradiated compared with normal DC hosts. The cytokines detected in the DC may at least in part stem from inflammatory cells accumulating around the chambers. This animal model should be useful in further investigation of the highly complex regulatory network governing formation of white blood cells in the intact organism.

Animals↗
PubMed1989

Thymus and T cells are not essential for rat leucopoiesis.

It is generally held that T lymphocytes take part in the regulation of haemopoiesis. We have examined whether, in vivo, the influence of thymus of functional T cells is dispensable for the steady-state or accelerated formation of granulocytes (and in some experiments macrophages), utilizing a rat model. Untreated normal and athymic 'nude' rats had similar blood and marrow granulocyte counts and marrow proliferative activity. Bone marrow regeneration after two cytotoxic insults to the two kinds of rats gave no clues to an important regulatory role for thymic factors or T cells. Nor were such clues obtained in experiments where bone marrow cells from normal rats were cultured in vivo in diffusion chambers (DC), in either normal rats or rats undergoing a graft-versus-host (GvH) reaction (with supranormal serum levels of cytokines). On the other hand, when marrow cells from athymic rats were similarly cultured in DC, small but significant differences in leucopoiesis occurred between the three kinds of DC hosts: Marrow cells lacking functional T cells generated fewer proliferative granulocytes and had a lower proliferative activity when cultured in athymic than in normal hosts, whereas the proliferative granulocytes were most numerous in chambers carried by GvH rats. No differences were found for macrophages and eosinophilic granulocytes. The results indicate that thymic hormones or T cells or both can stimulate granulopoiesis. They apparently play no indispensable role in short-term leucopoiesis, however, since their influence was weak in our experimental models. Consequently, marked effects seen in vitro in simplified cellular systems may lose some importance in the more complex in vivo setting.

Animals↗
PubMed1988

In vivo evaluation of radiolabelled antibodies with antigen-coated polymer particles in diffusion chambers.

A method for the evaluation of in vivo immunolocalization of labelled monoclonal antibodies (MoAb) is presented. The technique is an alternative to the nude mouse xenograft system. The antigen reservoir is an intraperitoneal diffusion chamber (DC) filled with a suspension of antigen-coated polymer particles. Intravenously injected 125I-labelled MoAb are allowed to specifically bind to this artificial abdominal 'tumour', which can be removed and measured for radioactivity after animal killing. The model can be used as a preclinical in vivo method for the evaluation of labelled MoAbs prepared for immunodiagnostics or therapy. The DC system permits the amounts of both antibody and antigen to be controlled and antibody access to the antigen within the DC is presumably the same in every animal. The model permits a systematic comparison of different antibodies and antibody fragments, labels and labelling procedures, as well as routes of administration in immunocompetent animals.

Alkaline Phosphatase↗
PubMed1988

Whole body distribution of 99mTc labelled autologous human granulocytes and radiation dose to cells and organs.

The whole body distribution of radioactivity as a function of time after infusion of 99mTc labelled autologous granulocytes was measured in three volunteers by means of a scanning bed, a scintillation camera and a minicomputer. Labelling was performed with a bisalt method without pretinning. There was a considerable initial lung sequestration (22%-31%) of the injected activity, which disappeared with an effective half life of 42 min. One h after infusion the activity was found mainly in the liver (41%), spleen (8%), lungs (9%) and kidneys (5%). Urine excretion amounted to 30% during the first 32 h after infusion. An injected activity of 100 MBq caused a radiation dose of 4.4 m Gy to the liver, 6.3 m Gy to the spleen, 3.7 m Gy to the kidneys, and 0.2 m Gy and 0.1 m Gy to the ovaries and testes respectively. The labelling procedure and the subsequent decay within the granulocytes gave them an absorbed radiation dose of 1.8 Gy after 25 min (i.e., at completion of the infusion) and 8.4 Gy after 4 h (i.e., the normal imaging time). In vitro tests revealed no signs of radiation damage to the cells.

Adult↗
PubMed1988

Interferons affect oxygen metabolism in human neutrophil granulocytes.

Human polymorphonuclear neutrophil granulocytes (PMN) were incubated with recombinant interferons (IFNs) and tested for O2 consumption, hydrogen peroxide formation, and chemiluminescence. N-formyl-methionyl-leucyl-phenylalanine (f-MLP, a bacterial peptide analogue) and phorbol myristate acetate (PMA, a protein kinase C activator) were used as PMN stimuli. An increase in O2 consumption after f-MLP-stimulation was seen when PMN had been incubated 2-4 h with either 1000 IU/ml IFN-alpha or 100 IU/ml IFN-gamma, but this increase in O2 consumption was not observed with 1000 IU/ml IFN-beta. Likewise, 100 U/ml IFN-gamma enhanced f-MLP stimulated chemiluminescence, whereas IFN-alpha or IFN-beta (1000 U/ml) had no detectable effects. None of the interferons affected baseline or PMA-stimulated O2 consumption and chemiluminescence, nor did they influence the H2O2-dependent oxidation of intracellular dichlorofluorescein (DCFH) (baseline, f-MLP-stimulated or PMA-stimulated). Our data indicate that some--but not all--aspects of oxygen metabolism in PMN can be affected by IFN, and that there are differences between various subtypes of IFNs regarding their neutrophil priming potential.

Granulocytes↗
PubMed1988

Dual effect of thiopentone on human granulocyte activation. Non-intervention by ketamine and morphine.

The immune system, defending our organism against infections, can also cause disease. Anaesthetics may impair immunological defence by modifying the number and functions of immunocompetent cells, including the polymorphonuclear leucocytes (PMN). We have studied the effects of thiopentone, ketamine and morphine on some stimulated PMN responses that presumably reflect their microbicidal activity, i.e. oxygen consumption, aggregation, and volume increase. Stimulators were N-formyl-methionyl-leucyl-phenylalanine (FMLP, affecting cells via specific membrane receptors) and phorbol-myristate-acetate (PMA, activating protein kinase C, thereby short-cutting intramembraneous steps in normal signal transmission, and presumably provoking near-maximal cell responses with the dose applied). Preincubation of PMN with low doses of thiopentone enhanced oxygen consumption in unstimulated cells as well as in response to FMLP, but not PMA. FMLP-stimulated volume and aggregation responses were not detectably affected. The highest concentration of thiopentone depressed both oxygen uptake and volume/aggregation responses in FMLP-stimulated PMN. The amount of oxygen consumed after PMA stimulation was not affected, but both the onset of increased consumption and the maximal response were delayed. The two other drugs investigated, ketamine and morphine, did not appreciably affect oxygen consumption or aggregation by PMN: neither the baseline values nor those obtained after FMLP or PMA stimulation.

Cell Aggregation↗